10 research outputs found
Autoinduced AND Gate Controls Metabolic Pathway Dynamically in Response to Microbial Communities and Cell Physiological State
Quorum
sensing (QS) systems have been widely applied in biotechnology
and synthetic biology that require coordinated, community-level behaviors.
Meanwhile, the cell physiological state is another key parameter that
affects metabolic pathway regulation. Here, we designed an autoinduced
AND gate that responds to both microbial communities and the cell
physiological state. A series of tunable QS systems in response to
different cell densities were obtained through random mutagenesis
of LuxR and optimization of the <i>luxRI</i> promoter; the
corresponding suitable stationary phase sensing system was selected
after monitoring the fluorescence process during cell growth. The
application of the final synthetic device was demonstrated using the
polyhydroxybutyrate (PHB) production system. The AND gate system increased
PHB production by 1–2-fold in <i>Escherichia coli</i>. This synthetic logic gate is a tool for developing a general dynamic
regulation system in metabolic engineering in response to complex
signals, without using a specific sensor
MOESM1 of Easy regulation of metabolic flux in Escherichia coli using an endogenous type I-E CRISPR-Cas system
Additional file 1. All strain’s name, plasmids, primers used in the study as well as figures mentioned in the main text are available in this file
Dual-phase fermentation of YL104H in glucose, xylose, or mixed glucose–xylose.
<p>Dual-phase fermentation of YL104H in glucose, xylose, or mixed glucose–xylose.</p
ATP-Based Ratio Regulation of Glucose and Xylose Improved Succinate Production
<div><p>We previously engineered <i>E</i>. <i>coli</i> YL104H to efficiently produce succinate from glucose. Furthermore, the present study proved that YL104H could also co-utilize xylose and glucose for succinate production. However, anaerobic succinate accumulation using xylose as the sole carbon source failed, probably because of an insufficient supply of energy. By analyzing the ATP generation under anaerobic conditions in the presence of glucose or xylose, we indicated that succinate production was affected by the intracellular ATP level, which can be simply regulated by the substrate ratio of xylose to glucose. This finding was confirmed by succinate production using an artificial mixture containing different xylose to glucose ratios. Using xylose mother liquor, a waste containing both glucose and xylose derived from xylitol production, a final succinate titer of 61.66 g/L with an overall productivity of 0.95 g/L/h was achieved, indicating that the regulation of the intracellular ATP level may be a useful and efficient strategy for succinate production and can be extended to other anaerobic processes.</p></div
Succinate production in a 3-L bioreactor.
<p>Succinate production, sugar consumption and biomass of YL104H in artificial sugar mixture (A) and xylose mother liquor (B) under aerobic, microaerobic and anaerobic whole-phase fermentations. The xylose mother liquor contained approximately 300 g/L xylose and 100 g/L glucose, and 50 g/L glucose was added to adjust the glucose: xylose ratio at 1:2.</p
Succinate production in a 3-L bioreactor.
<p>Succinate production, sugar consumption and biomass of YL104H in artificial sugar mixture (A) and xylose mother liquor (B) under aerobic, microaerobic and anaerobic whole-phase fermentations. The xylose mother liquor contained approximately 300 g/L xylose and 100 g/L glucose, and 50 g/L glucose was added to adjust the glucose: xylose ratio at 1:2.</p
The anaerobic ATP production or consumption on different carbon sources predicted using flux balance analysis (FBA).
<p>Solid rectangles denote inactivated pathways, and the metabolic fluxes of reactions were stated alongside. The first, second and third row of numbers corresponding to glucose, mixed glucose-xylose (1:1) and xylose.</p
Relationship among succinate production, sugar ratio and intracellular ATP level.
<p>The ATP level was shown as the slops of their ATP regression lines which represented the specific rate of ATP consumption in units of nmol/(g CDW)/h. The solid and dashed lines were the second order polynomial regression line calculated by Origin Pro 8.0.</p
Intracellular ATP levels in the presence of different carbon sources.
<p>Data are the ATP contents per cell dry weight, and the regression lines were calculated by Origin Pro 8.0. All slopes (<i>k</i>) of these regression lines were listed beside, and their regression coefficients were all above 0.95. All the date were measuring at the anaerobic fermentation stage between 28 to 70 hours.</p
Dual-phase fermentation of YL104H for different glucose-to-xylose mass ratios.
<p>Dual-phase fermentation of YL104H for different glucose-to-xylose mass ratios.</p